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Production of recombinant EGFP via surface display of ice nucleation protein and self-cleavage intein

Paper ID Volume ID Publish Year Pages File Format Full-Text
3789 189 2011 6 PDF Available
Title
Production of recombinant EGFP via surface display of ice nucleation protein and self-cleavage intein
Abstract

In this study, a novel recombinant protein production system was developed by constructing a protein chimera with a self-cleavage segment along with a cell surface display segment to finally produce a model protein entity (i.e., enhanced green fluorescent protein, EGFP). In the plasmid construction, the EGFP gene was fused to genes of a self-cleaving intein (INT) and an ice nucleation protein (INP) which can anchor on the cell membrane. In the cultivation of recombinant Escherichia coli, the cells expressed high green florescence after the addition of isopropyl-β-d-thiogalactopyranoside (IPTG) as the inducer. By simply holding the cell pellets in Tris–HCl (pH 10.0) buffer at room temperature, EGFP was solubilized from the INP–INT segment embedded on the cell surface via intein's self-cleavage function. The EGFP concentration of 273 mg/L and recovery of 88% were obtained at day 5, whereas the best EGFP productivity of 187 mg/L/d was obtained at day 1. The EGFP can be harvested only via centrifugation, and no cell disruption process is required. This simplified approach is expected to be applicable for obtaining recombinant functional proteins for academic and industrial use.

Graphical abstractFigure optionsDownload full-size imageDownload as PowerPoint slideHighlights► In this study we develop a novel recombinant EGFP production approach. ► We construct a plasmid with INP, INT and the EGFP genes. ► The protein chimera of INP–INT–EGFP can be induced in Escherichia coli cell surface. ► By holding the cell pellets in Tris-buffer, EGFP is released from cell surface. ► EGFP is harvested via centrifugation, no cell disruption is needed.

Keywords
Recombinant protein production; Surface display; Ice nucleation protein; Intein; Enhanced green fluorescent protein; Protein purification
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Production of recombinant EGFP via surface display of ice nucleation protein and self-cleavage intein
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Publisher
Database: Elsevier - ScienceDirect
Journal: Biochemical Engineering Journal - Volume 54, Issue 3, 15 May 2011, Pages 158–163
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us